Frequency of Intestinal Parasites among Zoo Animal by Morphometric Criteria and First Report of the Bivitellobilharzia nairi from Elephant (Elephasmaximus maximus) in Iran.

Background
Intestinal parasitic infections are major causative agents of wildlife health complications among different parts of the world. This study aimed to investigate the gastro-intestinal parasites in feces of the zoo animals based on parasitological and morphometric criteria.


Methods
One hundred fresh fecal samples were collected from 35 species of animal lived in Eram park zoo, Tehran, Central Iran during Oct 2015 to Jun 2015. All collected samples were examined by microscopic observation following direct wet mount preparation (normal saline and Lugol's iodine), formalin-ether concentration, and permanent staining. The morphometric aspects of the recovered eggs were surveyed with the aid of Camera Lucida (×400).


Results
65.7% (23/35) of zoo animal species were infected with intestinal parasites. The superfamily Trichostrongyloidea (6/16) and Strongylus sp. (16/4) were the most prevalent helminthic infections, while Blastocystis sp. (6/14), Entamoeba cyst (3/14) and Eimeria sp. (3/14) were the common protozoan parasites. For the first time, Bivitellobilharzia nairi egg was identified an elephant at Iran. Intestinal parasitic infections were apparently circulating among animals of the Eram park zoo.


Conclusion
Identified parasitic infections can consider as a threatening source to visitors and workers' health that have contact with animals or their feces. Therefore, the effectual preventive strategies should be addressed to determine the risk factors, mechanisms of cross-transmission of parasite, the importance of applying the hygienic practices and well adjusting deworming programs for the animals, zoo workers and visitors.

Introduction nimals (zoo animal, wild animal, and domestics) have important role as host and reservoir of many zoonosis parasites. Zoological parks exhibit wild animals (mammals, birds, reptiles, amphibians, fish, etc.,) for aesthetic, educational, and conservation purposes (1). However, the intestinal parasitic diseases are one of the major health concerns causing illness and even mortality among zoo animals while in captivity, the effects of which range from subclinical to death (2). Parasitic diseases play a principal role in animal health ranging from negative impacts on the host population size to the evolution of host behaviors. Parasitic infections constitute one of the main challenges in wild animals in captivity (3)(4)(5). Insufficient data about parasitic contaminations among zoo animals in Iran is a major limiting issue in zoological gardens. Identification of parasitic infections is important to study the prevalence, geographical distribution, taxonomic status and biology of parasites (6). There is no hesitation that a regular and consistent program of gastrointestinal parasite investigation and measures of control based on accurate diagnosis, effective treatment and suitable prophylaxis would surely assist in reversing the condition of ill health in zoo animals. By trying to establish a profile of intestinal parasites in the zoo animals, valuable data will be gained for the development of public health and preventive remedy. Evaluation of intestinal parasites in zoo animals and different geographic regions has medical and veterinary importance to prevent transmission of intestinal parasitic diseases to human and domestic animals. Many studies have been documented the intestinal parasites of the zoo animals in different populations of the world (1,2,(5)(6)(7)(8).
Schistosomatidae families are helminthes with worldwide distribution, almost lived in all temperate condition and tropical regions. Schistosoma nairi (Bivitellobilharzia nairi) is a blood fluke (the blood vessels of the liver parenchyma) of Asian and African elephants (namely B. loxodontae) and usually remains subclinical. The first report of B. nairi reported in 1955 and named it as S. nairi. Subsequently occurrence of B. nairi was reported in elephants of India. In addition, in Sri Lanka and other region demonstrated the S. nairi (7,(9)(10)(11)(12).
There is no comprehensive research on the identification and prevalence rates of intestinal parasites among Iranian zoo animal. This study was designed to determine the parasitic infections among zoo animals based on parasitological and morphometric criteria.

Sample collection and morphometric studies
One hundred fresh fecal samples were collected from 35 species of animal lived in Eram A park zoo in Tehran and were placed into sterile sample bottles. Collection of fecal samples was carried out by the assistance of the animal handlers. Samples were stored at 70% ethanol alcohol and transported to the Medical Parasitological Laboratory of Shahid Beheshti University of Medical Science, Tehran, Iran. Direct wet mount (normal saline and Lugol's iodine) preparation, formalin-ethyl concentration technique, and permanent stains such as Trichrome and Ziehl Neelsen staining were examined for all suspected samples. The morphometric aspects of isolated eggs were made with the aid of Camera Lucida (×400).

Discussion
In this study, the various frequency rates of intestinal protozoan and helminthic infections were explicitly isolated and characterized by parasitological and morphometric investigations in Eram park zoo, Tehran where there is no comprehensive study yet.
The majority of the animals studied in this garden were infected with at least one intestinal parasite species. In this study, 65.7% of animal species (mammals, birds, reptiles, amphibians, fish, and etc.) including eight species of helminths (Trichostrongyloidea, Trichuris sp., Strongylus sp., Capillaria sp., Dictyocaulus, Dipylidium (gravid proglottid), Toxocara and B. nairi) and six species of protozoan parasites (Blastocystis sp., Iodamoeba, Endolimax nana, Cryptosporidium sp., Entamoeba sp. and Eimeria sp.) were infected by intestinal parasites. This may be due to suitable conditions of life cycle (temperature, humidity, intermediate hosts, and paratenic hosts., etc.,) of this area, provided the development of intestinal parasites.  (13). Animals are infected with various ranges of gastrointestinal parasites, which are representatives of the important pathogenic parasites found in equids worldwide. In the other surveys (Nigeria, Sri Lanka, African Republic, Nepal and Lithuania) some of the intestinal parasites observed in this study have also been reported (1-4, 6-8, 11,12). Most intestinal parasites observed in these animals can cause intestinal parasitic complications (including; anorexia, vomiting, diarrhea, anemia, loss of weights, etc.,) in humans, particularly animal handlers and visitors (14)(15)(16)(17)(18).
In this study, the B. nairi egg was detected for the first time from an elephant in Iran. The scanning electron microscope showed the tegument surface of male B. nairi is smoother than in other schistosomes. Furthermore, the B. nairi egg is similar morphologically to other Schistosoma spp, including; S. hippopotami, Ornithobilharzia dattai, and O. harinasutai (12).
There was no significant different among protozoa and helminth prevalence in this garden. In a zoological garden at the Kenya occurrence of helminths (64.4%) reported lower than protozoa (17.1%) and is otherwise with our study (19). In Belgian zoological gardens, the frequency of intestinal nematodes (36.5%) was also identified among captive primates (20).
Among the intestinal parasitic infection, the Trichostrongyloidea and Strongylus were the most prevalent helminth infections and Blastocystis sp., Entamoeba cyst and Eimeria sp. were observed in every group of animals studied. All of the parasites observed are zoonosis intestinal parasites and may be gastrointestinal disorders among human (zookeepers and possibility to visitors) and animals.
Zisitors, and zoo officials are exposure to direct contact with the excreta of these animals, therefore, to make sound decisions on regular deworming as well; using improved anthelmintic treatment guidelines should be broadly revised and addressed among the zoo animals and even zookeepers. Employing molecular methods should be employed to identify the accurate species of parasites.

Conclusion
The high prevalence rate of identified parasitic infections can consider as a economical loss of natural number of zoo animals and threatening source to visitors and workers' health that have contact with animals or their feces. Therefore, the effective preventive strategies should be considered to establish the risk factors, mechanisms of crosstransmission of parasite, the importance of employing the hygienic practices and well adjusting deworming programs for the animals, zoo workers and visitors.